The aim of this study was to investigate the effects of recombinant human-soluble thrombomodulin (rTM) on lipopolysaccharide (LPS)-induced, platelet-dependent neutrophil extracellular trap (NET) formation (NETosis). Human peripheral blood neutrophils and platelets were co-incubated with or without LPS (0.2 μg/ml) in the presence and absence of rTM (2 μg/ml). NETosis was confirmed by immunostaining and confocal microscopy. In the absence of platelets, LPS did not induce NETosis in the neutrophils. NETosis, however, was induced by LPS when neutrophils were co-cultured with platelets (64 % of neutrophils). Notably, rTM was able to fully inhibit NETosis in neutrophils cultured with platelets and in the presence of LPS. rTM did not induce NETosis in this co-culture system (p < 0.01 versus LPS in the absence of rTM). These results show that rTM can suppress LPS-induced platelet-dependent NETosis in vitro.

Neutrophil extracellular trap (NET) formation, an innate immune system response, is associated with thrombogenesis and vascular endothelial injury. Circulatory disorders due to microvascular thrombogenesis are one of the principal causes of organ damage. NET formation in organs contributes to the exacerbation of sepsis, which is defined as a life-threatening organ dysfunction caused by a dysregulated host response to infection. We have previously reported that recombinant human soluble thrombomodulin (rTM) reduces lipopolysaccharide (LPS)-induced NET formation in vitro. Here, we aimed to show that thrombomodulin (TM)-mediated suppression of NET formation protects against organ damage in sepsis. Mice were injected intraperitoneally (i.p.) with 10 mg/kg LPS. rTM (6 mg/kg/day) or saline was administered i.p. 1 h after LPS injection. In the LPS-induced murine septic shock model, extracellular histones, which are components of NETs, were observed in the liver and lungs. In addition, the serum cytokine (interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), macrophage chemotactic protein-1 (MCP-1), and interleukin-10 (IL-10)) levels were increased. The administration of rTM in this model prevented NET formation in the organs and suppressed the increase in the levels of all cytokines except IL-1β. Furthermore, the survival rate improved. We provide a novel role of TM in treating inflammation and NETs in organs during sepsis.

We investigated a possible alteration in the ability of neutrophils to produce active oxygen species in streptozotocin (STZ)-induced diabetic rats. The production of Superoxide of the neutrophils was assessed by luminol-dependent chemiluminescence (LDCL) after the stimulation by opsonized zymosan. Four days after STZ (60 mg/kg, ip) injection, blood glucose level increased by 399 ± 9 mg/dl and the LDCL activity was significantly reduced in diabetic rats (control group: 5.12±1.53 KC/min²/10⁶ cells, STZ group: 1.10±0.07 KC/ min²/10⁶ cells, p<0.01). At Day 17, blood glucose level was maintained high (598 ±8 mg/dl) and the LDCL activity (1.01 ± 0.39 KC/min²/10⁶ cells) was almost at the same level as that of diabetic rats at Day 4. Subcutaneous injection of insulin for 10 consecutive days increased the LDCL activity of diabetic rats in a dose-dependent manner (vehicle: 0.82±0.27 KC/ min²/10⁶ cells, 0.04 U/day: 1.61 ±0.09 KC/min²/10⁶ cells, 0.40 U/day: 1.99±0.47 KC/min²/10⁶ cells, 4.00 U/day: 3.33 ±0.43 KC/min²/10⁶ cells). The data obtained herein indicate that an increased susceptibility to bacterial infection in diabetic rats results from impaired neutrophil function to produce active oxygen species.

ABO blood groups have been implicated as potential risk factors for various diseases. However, no study has investigated the association between sepsis mortality and ABO blood types. We aimed to evaluate the impact of these blood types on mortality in patients with sepsis and septic shock. This retrospective observational study was conducted at two general hospitals in Japan. Patients diagnosed with sepsis or septic shock were included and divided into four groups based on blood type (O, A, B, and AB). The association between type O vs. other types and 28- and 90-day mortalities was evaluated using multivariate logistic regression analysis adjusted for age, sex, and Sequential (Sepsis-related) Organ Failure Assessment score. This study included 415 patients, of whom 131 (31.6%), 171 (41.2%), 81 (19.5%), and 32 (7.7%) had type O, A, B, and AB, respectively. Blood type O was not associated with 28-day (odds ratio: 1.7 p = 0.08) or 90-day mortality (odds ratio: 1.53, p = 0.091). However, type O was significantly associated with higher 90-day mortality (odds ratio: 3.26, p = 0.009) in patients with septic shock. The role of ABO blood type in risk stratification for septic shock and the mechanisms that potentially affect the prognosis of sepsis patients need further investigation.